Group B streptococcus is the predominant etiological agent of neonatal bacteremia, sepsis and meningitis, and has been correlated with preterm membrane rupture and premature birth. While a limited number of GBS virulence determinants have been identified, the contribution of iron and iron acquisition to the pathogenesis of GBS infection is unknown. All pathogens require iron and a correlation between bacterial virulence and iron acquisition has been established. However, as the concentration of available iron in the human host (10 [-18]M) is well below that required by bacteria (10 [-8] M), successful pathogens express specific systems to acquire sufficient concentrations of intracellular iron. One such system involved in siderophore-mediated iron uptake, is the focus of this application. Siderophores are high-affinity iron chelators secreted from the bacterial cell to scavenge iron from host iron-binding proteins. Specialized uptake systems transport the siderophore-iron complex across the bacterial membrane. We have identified a putative siderophore-mediated iron transport system, the fhu operon, in GBS. The operon is comprised of four genes, fhuC, fhuD, fhuB, and fhuG, encoding a putative ATP-hydrolysis protein, siderophore (ferrichrome)- iron receptor protein, and two permeases, respectively. In this application, the role of the fhu operon in iron acquisition by GBS will be examined. Basic information on the requirement of GBS for iron, the ability of GBS to utilize siderophores as an iron source, and whether the organism secretes siderophores to acquire iron will first be established. The biochemical characterization of an isogenic mutant strain deficient for fhu will examine the role of this operon in GBS siderophore-iron transport. In order to define the siderophore specificity of the fhu operon, the construction and biochemical characterization of isogenic mutant strains deficient for each gene is proposed. In addition, heterologous expression studies will provide further evidence for the role of the fhu-encoded proteins in siderophore-iron transport. These studies will provide a basic understanding of the molecular mechanism of iron acquisition in GBS and will allow us to design appropriate in vivo studies to determine the role of iron transport in GBS pathogenesis. The potential of identifying novel therapeutic targets to prevent or treat infections is also proposed.